Abundant evidence indicates a genetic contribution to congenital heart disease (CHD), but the specific genes responsible for most CHD are still unknown. Previously identified CHD mutations have been found in family-based investigations. However, most CHD arises as an unexpected and unexplained event, likely due to incomplete penetrance of some gene variants and impaired reproductive fitness associated with others. This is particularly true for severe CHD which, if untreated, causes early lethality. In this application, we propose to focus on sporadic rather than familial CHD. Subjects will have severe CHD defined as cyanotic lesions and others that usually require intervention in infancy. Based on evidence that abnormal dosage of critical developmental genes causes some CHD, we will conduct genome-wide analyses for de novo copy number variants in trios of the affected child and unaffected parents. We will also assess alleles that contribute to severe CHD through case-control whole genome association studies. We will determine whether somatic mutations account for some sporadic, severe CHD and will analyze gene transcription in human malformed hearts. Together our studies will assess the contribution of previously defined CHD genes, identify new loci, genes, mutations and mechanisms, and evaluate genotype-phenotype relationships. Our studies harness modern genomic approaches and will use novel sequencing and transcription profiling strategies. We anticipate these investigations will foster highly productive collaborations with other research centers in this Consortium and will contribute fundamental knowledge about human cardiogenesis to the Cardiac Development Consortium. We propose four specific aims: 1. Use subgenome capture strategies to interrogate the sequence and copy number of known and candidate CHD genes, as well as newly identified CHD genes, and assess genotype-phenotype correlations. 2. Determine if somatic mutations cause or modulate CHD. 3. Define novel severe CHD loci (by discovery of de novo copy number variants and by genome-wide association studies), genes, and mutations. 4. Define RNA expression in affected regions of malformed human hearts. Three cores will support these studies: an Imaging Core for Complex CHD Phenotyping, a Genomic Analyses Core, and a Sequence Analysis Core. RELEVANCE (See instructions): Our studies will define new genes and mutations that cause severe CHD, enable genotype-phenotype studies, provide a framework to study the relationship of genetic factors to long-term outcomes, and provide data on recurrence risks. Data from the proposed studies, as well as our faculty, clinical and genetics resources, will advance the goals of the NHLBl's Pediatric Cardiovascular Translation Consortium.